Type 1 diabetes (T1D) is a very serious autoimmune disease resulting from self- destruction of the insulin producing pancreatic ? cells. A disorder without a cure, T1D has seen a pronounced increase in its frequency over the last half-century at a global level. Despite advances in disease management, it remains a disorder that instills major morbidity (e.g., blindness, kidney and heart disease, risk for hypoglycemia) and mortality challenges on those afflicted. We portend a major reason that the community of T1D researchers has seen limited progress, over many decades, in attempts to identify therapies capable of preventing and/or reversing the disease is tied to a collective lack in understanding the mechanisms of disease development in humans; especially in terms of knowing the specific contributions of islet endocrine cells and the immune system to the disorder, including and especially their interplay with each other. This application proposes to test the hypothesis that a highly innovative technique known as High Multiplexed Imaging (HMI), that when applied to rare but informative pancreatic tissues from those with or at varying levels of risk for T1D as well as control tissues, will unlock research mysteries surrounding the disorder that have existed for many years. That hypothesis will be tested in this proposal by the performance of two interactive specific aims, the first being to establish HMI as a tool for pancreatic tissue analysis, while the second involves characterization of islet and immune system cells in T1D by HMI using validated antibodies. Importantly, to achieve said goals with maximal probability for success, we have formed a team well poised to address the lofty challenges associated with our proposal. This proposal combines the efforts of three well-established investigators that developed HMI (Bodenmiller), have provided crucial information regarding islet cell plasticity (Herrera), and helped define the natural history of T1D in both humans and the NOD mouse model of the disease (Atkinson). Our team is committed to not only generate information vital for our own proposed project goals but in addition, to generating data for the entire Human Islet Research Network (HIRN) collective. In sum, we firmly believe this study will result in a technological means for providing novel information regarding the pathogenesis of T1D; knowledge that should prove beneficial for attempts seeking to prevent and/or reverse the disorder.